The objective of this project is to probe similarities and differences in protein active site topography, bound substrate topography, and mechanism of hydrolytic action among serine proteases, especially bacterial subtilisins and vertebrate chymotrypsins, using chemical methods. Active site and complementary substrate topographies are being studied using structure-reactivity relationships obtained with conformationally rigid models, esters of (S)-2-acylamido-1,2,3,4-tetrahydro-2-naphthoic acids and related compounds. The phenyl, alpha-acylamino, and ester functions of these substrates are locked in a stereorelationship identical to that found x-ray crystallographically for (S)-acylphenylalanine bound at the active site of subtilisin BPN' and gamma-chymotrypsin. A good rigid model for the reactive conformation of specific substrates should elicit in their enzyme-catalyzed hydrolyses the same response to substrate structural modification as the same modification in the natural substrate. The effect of modification of the side chain phenyl, alpha acylamino, and ester leaving group in the natural and model substrates on reactivity, stereoselectivity, and mechanism are being explored using pH dependences, deuterium isotope effects, and linear free energy relationships. These studies may reveal details of the degree of development of similarity in the catalytic machinery of the subtilisins and chymotrypsins and provide new insights into the concept of convergent evolution. BIBLIOGRAPHIC REFERENCE: "Acylation of Subtilisin Carlsberg by Phenyl Esters", M.S. Matta, C.M. Greene, R.L. Stein and P.A. Henderson, J. Biol. Chem., in press (Feb/Mar 1976 publication).